RS Medium Base (Rimler-Shotts Medium Base) M576

Intended use
Rimler-Shotts (RS) Medium Base is used for selective isolation, cultivation and presumptive identification of Aeromonas
hydrophila from clinical and non-clinical samples.
Composition**
Ingredients Gms / Litre
Yeast extract 3.000
Maltose 3.500
L-Cysteine hydrochloride 0.300
L-Lysine hydrochloride 5.000
L-Ornithine hydrochloride 6.500
Sodium thiosulphate 6.800
Ferric ammonium citrate 0.800
Sodium deoxycholate 1.000
Sodium chloride 5.000
Bromothymol blue 0.030
Agar 13.500
Final pH ( at 25°C) 7.0±0.2
**Formula adjusted, standardized to suit performance parameters
Directions
Suspend 45.43 grams in 990 ml purified / distilled water. Heat to boiling to dissolve the medium completely. DO NOT
AUTOCLAVE. Cool to 45-50°C and aseptically add rehydrated content of 1 vial of Novobiocin Supplement (FD096). Mix
well and pour into sterile Petri plates.
Principle And Interpretation
RS Medium was formulated by Rimler and Shotts (6) based on the principle of Xylose-Lysine (XL) Agars (7,8). It is used for
selective isolation and presumptive identification of Aeromonas hydrophila and other gram-negative bacteria based on their
ability to decarboxylate lysine and ornithine, maltose fermentation and H2S production (4).
Yeast extract acts as a source of nutrients. Sodium thiosulphate, L-cysteine hydrochloride and ferric ammonium citrate are
the indicators of H2S production. The medium contains maltose, which is mostly fermented by all Aeromonas. Maltose
fermentation is indicated by bromothymol blue. Sodium deoxycholate and novobiocin inhibit gram-positive bacteria and
Vibrio species. Citrobacter freundii usually produce H2S but occasionally negative strains exist. The medium contains L-
cysteine and L-ornithine, which are often decarboxylated by enteric bacteria to give alkaline products. Lysine positive and
ornithine positive strains of Aeromonas may not have the typical strong yellow colour because of alkaline products produced
during decarboxylation of the amino acids. Results are interpreted within 24 hours since after 26 hours slow reversion of
yellow colour to a basic (green) colour occurs. Medium should be incubated at 35°C, which will eliminate possible growth of
Aeromonas salmonicida , which may grow at reduced temperatures giving false-positive reaction. Test the yellow colonies
with or without black centers (of H2S) for oxidase to rule out Citrobacter species. Proteus mirabilis is inhibited on this
medium.

Type of specimen
Clinical samples - stool, Food samples ; Water samples.

Specimen Collection and Handling:

For clinical samples follow appropriate techniques for handling specimens as per established guidelines (2,3 ).
For food samples, follow appropriate techniques for sample collection and processing as per guidelines (5).
For water samples, follow appropriate techniques for sample collection, processing as per guidelines and local standards.(1)
After use, contaminated materials must be sterilized by autoclaving before discarding.

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Warning and Precautions

In Vitro diagnostic use. Read the label before opening the container. The media should be handled by
trained personnel only. Wear protective gloves/protective clothing/eye protection/ face protection. Follow good
microbiological lab practices while handling specimens and culture. Standard precautions as per established guidelines
should be followed while handling clinical specimens. Safety guidelines may be referred in individual safety data sheets.

Limitations :
1. Pure isolate must be used

2. Results are interpreted within 24 hours since after 26 hours slow reversion of yellow colour to a basic (green) colour occurs.

Performance and Evaluation

Performance of the medium is expected when used as per the direction on the label within the expiry period when stored at
recommended temmperature.

Quality Control
Appearance
Light yellow to light green homogeneous free flowing powder

Gelling
Firm, comparable with 1.35% Agar gel.
Colour and Clarity of prepared medium
Dark green coloured clear to slightly opalescent gel forms in Petri plates
Reaction
Reaction of 4.54% w/v aqueous solution at 25°C. pH : 7.0±0.2
pH
6.80-7.20
Cultural Response
Cultural characteristics observed with added Novobiocin Supplement(FD096) after an incubation at 35-37°C for 24 hours .

Organism Inoculum Growth Maltose Lysine/ H2S

(CFU) fermentation Ornithine
decarboxylation

Aeromonas hydrophila 50-100 good positive negative negative

ATCC 7966 (00063*) reaction, yellow reaction reaction
coloured
colonies
Citrobacter freundii ATCC 50-100 good negative variable positive,
8090 reaction reaction black centered
colonies
Escherichia coli ATCC 50-100 good negative variable negative
25922 (00013*) reaction reaction reaction
Proteus vulgaris ATCC 50-100 good positive negative positive,
13315 reaction, yellow reaction black centered
coloured colonies
colonies
Salmonella Typhi ATCC 50-100 good positive negative negative
6539 reaction, yellow reaction reaction
coloured
colonies
Key : * Corresponding WDCM numbers

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 HiMedia Laboratories Technical Data

Storage and Shelf Life

Store between 10-30°C in a tightly closed container and the prepared medium at 2-8°C. Use before expiry date on
the label. On opening, product should be properly stored dry, after tightly capping the bottle in order to prevent lump
formation due to the hygroscopic nature of the product. Improper storage of the product may lead to lump formation.
Store in dry ventilated area protected from extremes of temperature and sources of ignition Seal the container tightly
after use. Use before expiry date on the label.
Product performance is best if used within stated expiry period.

Disposal
User must ensure safe disposal by autoclaving and/or incineration of used or unusable preparations of this product. Follow
established laboratory procedures in disposing of infectious materials and material that comes into contact with clinical
sample must be decontaminated and disposed of in accordance with current laboratory techniques (2,3).
References
1. Baird R.B., Eaton A.D., and Rice E.W., (Eds.), 2015, Standard Methods for the Examination of Water and
Wastewater, 23rd ed., APHA, Washington, D.C.
2. Isenberg, H.D. Clinical Microbiology Procedures Handbook. 2nd Edition.
3. Jorgensen, J.H., Pfaller, M.A., Carroll, K.C., Funke, G., Landry, M.L., Richter, S.S and Warnock., D.W. (2015)
Manual of Clinical Microbiology, 11th Edition. Vol. 1.
4.MacFaddin J. F., 1985, Media for Isolation-Cultivation-Identification-Maintenance of Medical Bacteria, Vol. I,
Williams and Wilkins, Baltimore.
5. Salfinger Y., and Tortorello M.L. Fifth (Ed.), 2015, Compendium of Methods for the Microbiological Examination of
Foods, 5th Ed., American Public Health Association, Washington, D.C.
6.Shotts E. B. Jr. and Rimler R., 1973, Appl. Microbiol., 26(4):550.
7.Taylor W. I. and Harris B., 1965, Am. J. Clin. Pathol., 44:476.
8.Taylor W. I., 1965, Am. J. Clin. Pathol., 44:471.

Revision : 02 / 2019

In vitro diagnostic medical

IVD device

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30°C Storage temperature

10°C

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